Diarrhea causes an estimated 5 to 10 million deaths per year in the developing world, and is the third leading cause of pediatric hospital admissions in the United States. A significant percentage of acute cases of diarrhea cannot be diagnosed despite recent advances in the recognition of enteric pathogens and elucidation of their mechanisms of pathogenicity. While certain categories of Escherichia coli have been demonstrated to be important agents of diarrheal disease, other strains with distinctive phenotypes which do not fall into these categories are frequently associated with gastroenteritis in children in the absence of any recognizable pathogen. These strains possess the ability to agglutinate human erythrocytes in the presence of mannose, and show a distinctive diffuse pattern of adherence to HEp-2 tissue culture cells. These adhesive properties are rarely observed among strains of E. coli from healthy individuals. Adhesins are known to be virulence factors in E. coli strains of proven pathogenicity, and therefore may be of significance in these gastroenteritis-associated strains. The research seeks to characterize the organization and function of genes encoding the adhesin of a representative strain. The genes will be isolated by recombinant DNA techniques, and specific gene products will be analyzed in an E. coli minicell system. Gene function will be investigated by minicell analysis of transposon-mediated insertion mutants in adhesin expression. The nucleotide sequence of the adhesin structural gene will be determined and compared with previously characterized adhesins of E. coli. A hybridization probe derived from the isolated adhesin gene will be used to search for related sequences in other strains of E. coli. Isolates will also be examined with antisera raised against purified adhesin protein. Genetically and/or serologically unrelated adhesins mediating the characteristic phenotype of mannose-resistant hemagglutination and diffuse HEp-2 cell adherence will be identified, purified, and specific gene probes prepared. In this manner it is hoped to define a class of E. coli adhesins potentially significant in diarrheal disease. This significance will be confirmed by genetic and serological examination of diarrheal and other isolates of E. coli for the presence of this class of adhesin. The research will further the understanding of adhesins as virulence factors of E. coli and enhance the ability to diagnose acute infectious diarrhea.